Leishmania is a parasitic protozoa, which is the causative agent of Leishmaniasis. This disease is the cause of significant morbidity and mortality throughout the tropical area of the world. The development of effective treatments has been hampered by the difficulty in isolating virulence genes. The goal of this proposal is to develop a genetic tool which will enable us to easily isolate mutants. RNAi is becoming a powerftil tool for whole-genome functional analysis in other organisms. The ability to eliminate gene expression by targeting the homologous mRNA instead of the gene makes RNAi a potentially powerful technique in this asexual diploid organism. Therefore, it is the goal of this proposal to determine the elements necessary to eliminate gene expression by RNAi. The specific aims of this proposal will determine the region of the gene necessary, whether the dsRNA or ssRNA is required, the requirement of Leishmania processing signals and the level of RNA expression.